Tyrosine hydroxylase from rat pheochromocytoma will be purified to homogeneity and a monospecific antiserum to the enzyme will be produced in rabbits. The enzyme from rat adrenal and striatum, guinea-pig vas deferens and mouse neuroblastoma cells will also be purified and the physical and kinetic characteristics of each enzyme and the changes in the kinetic properties of each enzyme produced by the cyclic AMP-dependent protein phosphorylating system, phosphatidyl-L-serine, cations and other putative regulators of the enzyme will be compared. The molecular basis of the physiological activation of tyrosine hydroxylase consequent to nerve stimulation will be explored, with particular attention directed to enzyme phosphorylation, or phosphorylation of an activator protein, and the role of calcium. Protein kinases and phosphoprotein phosphatases from each tissue will be purified and characterized and their possible relevance to the activation and deactivation of tyrosine hydroxylase will be determined. The mechanism of induction of tyrosine hydroxylase in the adrenal of rats and in mouse neuroblastoma cells in culture will be investigated further, and the role of protein kinase and cyclic AMP in this process will be ascertained. The ontogenic development of sensitivity to putative allosteric modulators of tyrosine hydroxylase will be defined in striatum of rat fetuses and young animals. The ultrastructural localization of tyrosine hydroxylase will be determind by immunocytochemical techniques and changes in the ultrastructural distribution of the enzyme with nerve stimulation will be examined. Regulation of catecholamine release consequent to nerve stimulation from isolated, perfused spleens will be evaluated with particular emphasis on cyclic nucleotides, alpha and beta adrenoceptor, dopamine and cholinergic agonists and adenosine. The uptake of tyrosine into adrenergic neurons and the equilibration of precursor amino acids in this structure will be assessed. Amino acid aminotransferases and keto acids will be evaluated with regard to their effects on maintenance of available neurotransmitter stores during prolonged nerve stimulation.